Chapter 25 Lecture Notes - CEM252 at MSU Fall 1996

Chapter 25: Nucleic Acids

DNA

• DNA (deoxyribonucleic acid) encodes proteins that enable us to function
• DNA forms a double helix - a secondary structural unit

DNA Forms a Double Helix Due to Hydrogen Bonding

Purine & Pyrimidine Bases

Draw dashed lines to represent hydrogen bonds between base pairs.

Chapter 25 - Slide 4

Nucleosides and Nucleotides

• Nucleoside: Amine base on ribose or deoxyribose
  

Biological Importance of Nucleotides

Biological nucleotides include:

• DNA
• RNA
• ATP (adenosine triphosphate) - the biological energy storehouse
  
• Some anti-cancer drugs
• Some anti-HIV drugs Chapter 25 - Slide 6
  

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  Adenosine Triphosphate (ATP)

  Discuss with people around you why ATP is such a high-energy molecule.
  Chapter 25 - Slide 7
  

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  Nucleoside Numbering

  
  

  (refer also to figure 25.5)
  Chapter 25 - Slide 9
  

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  History Behind the Secondary Structure of DNA

  • Proposed in 1953 based on x-ray diffraction data and a number of attempts to propose a structure consistent with that data
  • Nobel Prize (1962) awarded to Watson, Crick, and Wilkins (Rosalind Franklin contributed to the work but died in 1958) for this research
  Chapter 25 - Slide 10
  

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  The Secondary Structure of DNA

  • Antiparallel Strands held together by hydrogen bonds between bases
  • Non-planar ribose and phosphate backbone coils structure into one of several helical conformations
  Chapter 25 - Slide 11
  

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  Antiparallel Secondary Structure

  
  Chapter 25 - Slide 12
  

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  B-DNA : Predominant Helical Conformation

  • Major Groove: 2.2 nm
  • Minor Groove: 1.2 nm

  Ribose: (C)2Õ-endo conformation
  
  movie
  Chapter 25 - Slide 13
  

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  A-DNA : Alternate Helical Conformation

  Ribose: (C)3Õ-endo conformation
  
  movie
  Chapter 25 - Slide 14
  

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  DNA Tertiary Structure

  Supercoiling (see figures pg. 1101, 1104)

  • In duplex circular DNA (plasmids)
    • Enzymes add or remove coils from the helix - local strain
    • Strain causes circular DNA to supercoil to distribute strain
  • Chromatin - coiling around histone proteins
    • Histones are rich in arginine and lysine (positively charged)
    • DNA coils around histones - this complex of DNA and histones is called chromatin
  Chapter 25 - Slide 15
  

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  Solid-Phase DNA Synthesis

  Why synthesize DNA and/or RNA?

  • Site-directed mutagenesis: studies of the effect of single amino acid replacements in proteins
  • Antisense nucleotide drug development efforts
  Chapter 25 - Slide 16
  

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  DNA/RNA Synthesis: Problems

  • Primary amines (present in adenine, guanine, cytosine) are nucleophilic and basic - must be protected
  • Hydroxyl groups (2Õ, 3Õ, 5Õ) must be protected
  • 3Õ and 5Õ hydroxyl groups must be deprotected at different steps in the synthesis
  Chapter 25 - Slide 17
  

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  DNA Synthesis: Amine protection

  Amines are commonly converted to less reactive amides:

  

  Discuss with neighbors: Why are amides less nucleophilic than amines? Chapter 25 - Slide 18
  

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  DNA Synthesis: Hydroxyl Protection

  5Õ-OH can be selectively protected with large groups such as 4,4Õ-dimethoxy trityl (DMTr):

  

  3Õ-OH treated with methyl N,N-diisopropylchlorophosphoramidite which becomes phosphate:

  
  Chapter 25 - Slide 19
  

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  DNA Synthesis: Solid Phase

  • Step 1: Anchor 3Õ-OH to solid support
  • Step 2: Remove 5Õ protecting group with acid
  • Step 3: Join next nucleotide - phosphite triester formed
  • Step 4: Oxidize phosphite to phosphate
  • Step 5: Repeat cycle as needed
  • Step 6: Remove final 5Õ protecting group
  • Step 7: Remove amine protection and cleave from support
  Chapter 25 - Slide 20
  

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  Last modified 1/6/96

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  Dr. Abby Parrill
  Department of Chemistry
  Michigan State University
  
  

  These pages may be downloaded and linked from other pages freely for academic and educational purposes. Questions, problems, and errors should be sent to parrill@argus.cem.msu.edu.